Macromolecular structures and cellular machineries are constructed under direction of genes. A proper collection of mutants should enable us to dissect the structures or mechanisms involved in neuronal development and function. The proposed study employs Drosophila mutants to study basic neuronal mechanisms and relationships between neuronal function and form during development. A recently developed cell culture system of Drosophila larval CNS allows direct morphological observation and physiological recording of developing neurons. We will continue our effort to characterize cultured larval neurons to gain baseline information for studies of neurobiological problems in this system. Neurons with distinct morphological patterns can be classified in cultures derived from various defined CNS regions. Staining by available antibodies and monoclonal antibodies raised against larval CNS will be used to refine the classification. The distinction and overlap of expression of genes controlling ion channel function will be examined among cell types. Particularly, the effects of several K+ channel mutations, of which differential expression has been suggested, will be studied by patch clamp methods To follow up on our preliminary findings, the influence of hyperexcitability on neurite outgrowth pattern among different types of cultured neurons will be further analyzed by using the K+ channel mutants. Mutations of shits gene affect membrane recycling, resulting in arrest of growth cone activity and neurite extension above 29 degrees C. Heat pulses will be applied to genetic mosaics containing a few shits sensory neurons to evaluate the effects of developmental delay and membrane recycling arrest on axonal navigation and terminal arborization and competition. The shits gene product will be identified by gel electrophoresis to study its biochemical properties and subcellular localization. To extend from this protein to other functionally related components in the molecular network controlling membrane recycling, we will isolate new mutants whose defects suppress or enhance the shits phenotype.